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Abstract

Tumors shed large numbers of cells into the vasculature. It is supposed that these cells give rise to metastases that are the major cause of death from cancer. However, the isolation and characterization of the so called circulating tumor cells (CTCs) is still challenging since cells are rarely to find among millions of normal leukocytes. The objective of this thesis was to characterize colorectal cancer (CRC) - associated CTCs and disseminated tumor cells (DTCs) from bone marrow samples. We studied CTC incidence in blood samples of CRC patients with the help of the FDA-cleared CellSearchTM system. We found that the number of patients with CTCs and the amount of CTCs was significantly correlated with the stage of disease. In addition, a significant higher rate of patients with CTCs in tumor-draining venous blood compared to the central venous blood was found. Furthermore, genomic analyses of single CTCs have been performed. This comprised the evaluation of typical CRC-associated mutations such as point mutations in TP53, BRAF, KRAS as well as the detection of microsatellite instability (MSI). Additionally, some CTCs were used to study global chromosomal aberrations by comparative genomic hybridization technique. Results revealed a remarkable genomic heterogeneity among the CTCs of single patients. Moreover, we detected several cases of genomic disparity among CTCs and the prevailing clone of matched cancer tissue. To evaluate differential gene expression that might enable tumor cell dissemination, we studied CTCs that were obtained from patients’ blood samples and from an othotopic mouse model of metastatic CRC. Gene expression profiles of CTCs and cell samples from cancer tissue were compared. The down-regulation of cell adhesion molecules such as E-cadherin might be involved in tumor cell dissemination. However, a significant epithelial-mesenchymal transition (EMT) in CTCs could not be confirmed. DTCs isolated from human bone marrow samples seem to adopt a dedifferentiated phenotype and lack expression of CK20 and CK19. Absence of EGFR and the proliferation marker Ki67 confirmed previous reports of dormancy in DTCs. With the present work we were able to demonstrate that the molecular characterization of single CTCs is feasible. However, further studies are required to increase the knowledge about the molecular traits of CTCs which might help to improve therapy and prognosis in CRC.